The Microscopy Core will support each of the projects with state-of-the- art electron microscopy, laser scanning confocal and digital fluorescence microscopy equipment, services, and support. These will include: 1. High Resolution Electron Microscopic Immunolocalization in Nuclei and Centrosomes. Immunogold staining will be done on whole mount preparations, on conventional ultrathin sections, and on resinless sections. The Core will embed, section, examine samples with investigators from each project, and prepare micrographs. Facilities for digitization of micrographs and for digital image processing will also be studied. Second antibodies, fixatives, fixatives, and sterile carbon-coated grids for whole mounts, and other reagents will be provided. 2. Localization of Proteins and Nucleic Acids by Confocal Microscopy. The Microscopy Core will provide access to a Leica TCS SP microscope and will train users in its use. An image processing workstation away from the confocal microscope will be available for all users. Training and assistance with digital processing will also be provided. 3. Localization of Proteins and Nucleic Acids by Confocal Microscopy. Each project will have access to the Digital Imaging Center at the University of Massachusetts Medical Center. This center provides digital fluorescence microscopes, performs deconvolution of image sets, provides sophisticated digital image processing on Silicon Graphic 02 workstations, and provides a variety of image output devices including dye sublimination printers and slide makers. 4. Localization of Fluorescently Tagged Molecules in Living Cells. The Microscope Core will assist users with GFP-fusion protein detection living cells by either confocal or conventional digital fluorescence microscopy. 5. Digital Micrograph Processing. The Microscopy Core will provide both hardware and software for digitization of electron micrographs, processing of digital micrograph sets, three-dimensional reconstructions and processing of confocal data sets, computer deconvolution of three- dimensional micrograph sets, and image output services. The Core will also train users in the use of these facilities and assist and advice them in their appropriate use. 6. Intranet Communication and Electronic Publishing. The Microscopy Core will facilitate communication between groups and the exchange of micrographs and electronic data between the Projects of this proposal. This will be done by providing an intranet Website for the posting of micrographs, image data, microscope scheduling information, and draft manuscripts. A WebSite accessible from the InterNet will also allow the posting of results and published papers, disseminating the results from this Program Project o the larger scientific community.